title: Targeted killing of Leishmania donovani in vivo and in vitro with amphotericin B attached to functionalized carbon nanotubes.
abstract: OBJECTIVE
  This study describes the antileishmanial efficacy of the novel drug formulation of amphotericin B (AmB) attached to functionalized carbon nanotubes (f-CNTs) and compares it with AmB.
  
  
  METHODS
  f-CNTs were prepared in a two-step chemical carboxylation and amidation process. The AmB was then attached to make f-CNT-AmB and its construction was confirmed by Fourier transform infrared (FTIR) spectroscopy and transmission electron microscopy (TEM). The cytotoxicity of the constructed compound, f-CNT-AmB, was assessed in vitro using the J774A.1 macrophage cell line and in vivo using healthy BALB/c mice. Antileishmanial activity of AmB and f-CNT-AmB was assessed in vitro using a macrophage (J774A.1 cell line) model of Leishmania donovani infection. Antileishmanial activity was assessed in vivo by comparing the parasite load of hamsters treated with a 5 day course of AmB, f-CNTs or f-CNT-AmB initiated at 30 days after infection with L. donovani parasites.
  
  
  RESULTS
  The FTIR spectroscopy and TEM data demonstrate the successful attachment of AmB to f-CNTs. The in vitro cytotoxicity of AmB, f-CNTs and f-CNT-AmB was measured by the cytotoxic concentration required to kill 50% of the cells: 0.48±0.06 μg/mL; 7.31±1.16 μg/mL; 0.66±0.17 μg/mL, respectively, in the J774A.1 cell line. The in vivo toxicity assessment of the compounds in BALB/c mice revealed no hepatic or renal toxicity. Against intracellular amastigotes the in vitro antileishmanial efficacy of f-CNT-AmB was significantly higher than that of AmB (IC50 0.00234±0.00075 μg/mL versus 0.03263±0.00123 μg/mL; P≤0.0001). The percentage inhibition of amastigote replication in hamsters treated with f-CNT-AmB was significantly more than that with AmB (89.85%±2.93% versus 68.97%±1.84%; P=0.0004).
  
  
  CONCLUSIONS
  The results of these experiments clearly demonstrate that f-CNT-AmB has significantly greater antileishmanial efficacy than AmB and had no significant cytotoxic effects.
authors.affiliation: Infectious Disease Research Laboratory, Department of Medicine, Institute of Medical Sciences, Banaras Hindu University, Varanasi 221 005, India.
authors.forename: Vijay Kumar
authors.initials: VK
authors.lastname: Prajapati
authors.forename: Kalpana
authors.initials: K
authors.lastname: Awasthi
authors.forename: Shalini
authors.initials: S
authors.lastname: Gautam
authors.forename: Thakur Prasad
authors.initials: TP
authors.lastname: Yadav
authors.forename: Madhukar
authors.initials: M
authors.lastname: Rai
authors.forename: Onkar Nath
authors.initials: ON
authors.lastname: Srivastava
authors.forename: Shyam
authors.initials: S
authors.lastname: Sundar
mesh_terms: D000666:Amphotericin B / Q000008:administration & dosage* / Q000494:pharmacology* / Q000633:toxicity; D000818:Animals; D000981:Antiprotozoal Agents / Q000008:administration & dosage* / Q000494:pharmacology* / Q000633:toxicity; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D005260:Female; D007893:Leishmania donovani / Q000187:drug effects*; D007898:Leishmaniasis, Visceral / Q000188:drug therapy / Q000469:parasitology; D008264:Macrophages / Q000187:drug effects; D008297:Male; D008647:Mesocricetus; D051379:Mice; D008807:Mice, Inbred BALB C; D037742:Nanotubes, Carbon
chemical_list: D000981:Antiprotozoal Agents; D037742:Nanotubes, Carbon; D000666:Amphotericin B